An important strategy for improving outcome after spinal cord injury (SCI) is to achieve axon regrowth across lesions to reach functional neural targets. Various molecules have the potential to foster axon regrowth but cannot pass the blood brain barrier and exhibit activity in many central nervous system (CNS) regions, necessitating local delivery to achieve efficacy while avoiding side effects. Prolonged but temporary delivery is needed. Clinically translatable methods for such delivery are lacking. Our goal is to develop functionalized diblock copolypeptide hydrogels (DCH) as fully synthetic biomaterials that that can easily and safely be injected into, and near, SCI lesions to provide depots for sustained local release of multiple molecules that manipulate local cells and stimulate axons to regrow into healthy tissue. Our previous works demonstrates DCH safety and efficacy to deliver growth factors that exert predictable effects over distances of several mm in CNS. New preliminary data show that: (1) DCH depots injected 2 days after SCI are able to simultaneously deliver multiple growth factors that stimulate substantive regrowth of both sensory and propriospinal fibers throughout the SCI lesion core. We find that these regrowing axons track along cells with newly upregulated laminin expression, and that regrowth can be blocked by simultaneous delivery of function-blocking antibodies that disrupt laminin-integrin binding. (2) When DCH delivery of multiple growth factors is combined with attenuation of glial scar by deletion of STAT3 in transgenic mice, axons regrow beyond the lesion core into the distal glial scar. New data also show DCH can deliver hydrophobic small molecules like JSI, which inhibits STAT3 and attenuates scar formation in a manner comparable to our transgenic mice. (3) When multiple DCH depots are placed into both the lesion core and distal healthy tissue, we find considerable axon regrowth into healthy tissue areas that contains viable NeuN-positive neurons. The work proposed will build on these preliminary findings and use DCH depots injected after SCI to simultaneously deliver different types of molecules (including multiple protein growth factors, antibodies and small hydrophobic molecules that manipulate gene expression) in order to: (i) manipulate cells in scar and lesion core to enable and support axon regrowth, (ii) directly stimulate and guide axon regrowth into, through and beyond lesions into healthy tissue, (iii) dissect cellular and molecular mechanisms that underlie the axon regrowth stimulated by different molecules or combinations of molecules, and (iv) test whether regrowing propriospinal neurons that reach healthy tissue are able to contact neurons there and are able to form relay connections that improve locomotor function. This DCH depot approach will provide a powerful tool for the experimental investigation of cellular and molecular mechanisms after SCI, and will facilitate the extensive trial and error testing needed to identify appropriate molecules for potential clinical translation. In addition, because DCH are fully synthetic biomaterials, there is also a realistic potential for clinical translation of DCH for use in SCI.